Integrin antagonists

• US 20020042368A1
• Filed: 02/23/2001
• Published: 04/11/2002
• Est. Priority Date: 02/25/2000
• Status: Active Grant

• Current Assignee: Immunex Corporation (Amgen Inc.)
• Original Assignee: Immunex Corporation (Amgen Inc.)
• Inventors: Fanslow, William C. III, Cerretti, Douglas P., Black, Roy A., Poindexter, Kurt M.
• Granted Patent: US 7,074,408 B2
• Time in Patent Office: Days
• US Class Current: 514/12
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61P 1/04 for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 17/06 Antipsoriatics
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 19/10 for osteoporosis
• CPC Class Codes: A61P 27/00 Drugs for disorders of the senses
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 27/06 Antiglaucoma agents or miotics
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/04 specific for metastasis
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/02 Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C07K 14/705 Receptors; Cell surface antigens; Cell surface determinants tumour specific antigens C07K14/4748
• CPC Class Codes: C12N 9/6489 Metalloendopeptidases (3.4.24)

Abstract

The present invention provides methods and compositions for inhibiting the biological activity of integrins, for inhibiting endothelial cell migration. and for inhibiting angiogenesis. In particular, the invention provides compositions comprising ADAM disintegrin domains and methods for using said compositions. In preferred embodiments the methods and compositions of the invention are used to inhibit angiogenesis and to treat diseases or conditions mediated by angiogenesis.

Claims

1. A method of antagonizing the binding of an integrin to its ligands comprising contacting a cell that expresses the integrin with an effective amount of an ADAM disintegrin domain polypeptide.

2. A method according to claim 1 wherein the ADAM disintegrin domain polypeptide is administered to a mammal in need of such treatment.

3. The method of claim 2 wherein the mammal is afflicted with a condition selected from the group consisting of ocular disorders, malignant and metastatic conditions, inflammatory diseases, osteoporosis and other conditions mediated by accelerated bone resorption, restenosis, inappropriate platelet activation, recruitment, or aggregation, thrombosis, or a condition requiring tissue repair or wound healing.

4. A method according to claim 2 wherein the method is a method of inhibiting angiogenesis in a mammal in need of such treatment, comprising administering to the mammal an inhibition-effective amount of an ADAM disintegrin domain polypeptide, wherein the disintegrin domain does not contain an RGD sequence.

5. The method of claim 2 wherein the ADAM disintegrin domain is in the form of a multimer.

6. The method of claim 5 wherein the multimer is a dimer or trimer.

7. The method of claim 5 wherein the multimer comprises an Fc polypeptide or a leucine zipper.

8. The method of claim 2 wherein the ADAM disintegrin domain is from a human ADAM.

9. The method of claim 8 wherein the ADAM disintegrin domain is from an ADAM selected from the group consisting of ADAM-8, ADAM-9, ADAM-10, ADAM-15, ADAM-17, ADAM-20, ADAM-21, ADAM-22, ADAM-23, and ADAM-29.

10. The method of claim 9 wherein the ADAM disintegrin domain is from ADAM-17, ADAM-20, or ADAM-23.

11. The method of claim 2 wherein the ADAM disintegrin domain polypeptide comprises an amino acid sequence selected from the group consisting of:

(a) amino acids 1-494 of SEQ ID NO;

2, amino acids 23-264 of SEQ ID NO;

2, amino acids 1-533 of SEQ ID NO;

4, amino acids 23-303 of SEQ ID NO;

4, amino acids 1-465 of SEQ ID NO;

6, amino acids 23-235 of SEQ ID NO;

6, amino acids 1-522 of SEQ ID NO;

8, amino acids 23-292 of SEQ ID NO;

8, amino acids 1-446 of SEQ ID NO;

10, amino acids 23-216 of SEQ ID NO;

10, amino acids 1-535 of SEQ ID NO;

12, amino acids 23-305 of SEQ ID NO;

12, amino acids 1-523 of SEQ ID NO;

14, amino acids 23-293 of SEQ ID NO;

14, amino acids 1-542 of SEQ ID NO;

16, amino acids 23-312 of SEQ ID NO;

16, amino acids 1-540 of SEQ ID NO;

18, amino acids 23-310 of SEQ ID NO;

18, amino acids 1-528 of SEQ ID NO;

22, amino acids 23-298 of SEQ ID NO;

22;

(b) fragments of the polypeptides of (a) wherein said fragments retain at least one ADAMdis activity; and

(c) fusion polypeptides comprising the polypeptides of (a) or (b), wherein said fusion polypeptides retain at least one ADAMdis activity,

wherein the ADAMdis activity is selected from the group consisting of integrin binding activity, inhibition of endothelial cell migration, and inhibition of angiogenesis.

12. The method of claim 11 wherein the ADAM disintegrin domain comprises an amino acid sequence selected from the group consisting of amino acids 34-91 of SEQ ID NO:

2, 34-92 of SEQ ID NO;

4, 34-99 of SEQ ID NO;

6, 34-92 of SEQ ID NO;

8, 34-93 of SEQ ID NO;

10, 34-91 of SEQ ID NO;

12, 34-91 of SEQ ID NO;

14, 34-92 of SEQ ID NO;

16, 34-91 of SEQ ID NO;

18, or 34-91 of SEQ ID NO;

22.

13. The method of claim 2 wherein the ADAM disintegrin domain polypeptide is a variant that is at least 70%, 80%, 90%, 95%, 98%, or 99% identical in amino acid sequence to a polypeptide selected from the group consisting of:

(a) amino acids 1-494 of SEQ ID NO;

2, amino acids 23-264 of SEQ ID NO;

2, amino acids 1-533 of SEQ ID NO;

4, amino acids 23-303 of SEQ ID NO;

4, amino acids 1-465 of SEQ ID NO;

6, amino acids 23-235 of SEQ ID NO;

6, amino acids 1-522 of SEQ ID NO;

8, amino acids 23-292 of SEQ ID NO;

8, amino acids 1-446 of SEQ ID NO;

10, amino acids 23-216 of SEQ ID NO;

10, amino acids 1-535 of SEQ ID NO;

12, amino acids 23-305 of SEQ ID NO;

12, amino acids 1-523 of SEQ ID NO;

14, amino acids 23-293 of SEQ ID NO;

14, amino acids 1-542 of SEQ ID NO;

16, amino acids 23-312 of SEQ ID NO;

16, amino acids 1-540 of SEQ ID NO;

18, amino acids 23-310 of SEQ ID NO;

18, amino acids 1-528 of SEQ ID NO;

22, amino acids 23-298 of SEQ ID NO;

22; and

(b) fragments of the polypeptides of (a),

wherein said variant polypeptide retains at least one ADAMdis activity,

wherein the ADAMdis activity is selected from the group consisting of integrin binding activity, inhibition of endothelial cell migration, and inhibition of angiogenesis.

14. The method of claim 2 wherein the ADAM disintegrin domain polypeptide is encoded by a nucleic acid comprising a sequence selected from the group consisting of:

(a) nucleotides 118-1599 of SEQ ID NO;

1, nucleotides 184-909 of SEQ ID NO;

1, nucleotides 46-1644 of SEQ ID NO;

3, nucleotides 112-954 of SEQ ID NO;

3, nucleotides 25-1419 of SEQ ID NO;

5, nucleotides 91-729 of SEQ ID NO;

5, nucleotides 41-1606 of SEQ ID NO;

7, nucleotides 107-916 of SEQ ID NO;

7, nucleotides 25-1362 of SEQ ID NO;

9, nucleotides 91-672 of SEQ ID NO;

9, nucleotides 25-1629 of SEQ ID NO;

11, nucleotides 91-939 of SEQ ID NO;

11, nucleotides 25-1593 of SEQ ID NO;

13, nucleotides 91-903 of SEQ ID NO;

1 3, nucleotides 25-1650 of SEQ ID NO;

15, nucleotides 91-960 of SEQ ID NO;

15, nucleotides 25-1644 of SEQ ID NO;

17, nucleotides 91-954 of SEQ ID NO;

17, nucleotides 118-1701 of SEQ ID NO;

21, nucleotides 184-1011 of SEQ ID NO;

21;

(b) sequences which, due to the degeneracy of the genetic code, encode a polypeptide encoded by a nucleic acid of (a); and

(c) sequences that hybridize under conditions of moderate or high stringency to a sequence of (a) or (b) and that encode a polypeptide that retains at least one ADAMdis activity,

wherein the ADAMdis activity is selected from the group consisting of integrin binding activity, inhibition of endothelial cell migration, and inhibition of angiogenesis.

15. The method of claim 11 wherein the ADAM disintegrin domain polypeptide has been produced by culturing a recombinant cell that encodes the ADAM disintegrin domain polypeptide under conditions permitting expression of the ADAM disintegrin domain polypeptide, and recovering the ADAM disintegrin domain polypeptide.

16. The method of claim 11 wherein the ADAM disintegrin domain polypeptide is present in a composition comprising a pharmaceutically acceptable carrier.

17. The method of claim 11 wherein the mammal has a disease or condition mediated by angiogenesis.

18. The method of claim 17 wherein the disease or condition is characterized by ocular neovascularization.

19. The method of claim 17 wherein the disease or condition is a solid tumor.

20. The method of claim 11 wherein the method further comprises treating the mammal with radiation.

21. The method of claim 11 wherein the method further comprises treating the mammal with a second therapeutic agent.

22. The method of claim 21 wherein the second therapeutic agent is selected from the group consisting of alkylating agents, antimetabolites, vinca alkaloids and other plant-derived chemotherapeutics, antitumor antibiotics, antitumor enzymes, topoisomerase inhibitors, platinum analogs, adrenocortical suppressants, hormones and antihormones, antibodies, immunotherapeutics, radiotherapeutics, and biological response modifiers.

23. The method of claim 21 wherein the second therapeutic agent is selected from the group consisting of cisplatin, cyclophosphamide, bleomycin, carboplatin, fluorouracil, 5-fluorouracil, 5-fluorodeoxyuridine, methotrexate, taxol, asparaginase, vincristine, vinblastine, mechloretamine, melphalan, 5-Iluorodeoxyuridine, lymphokines and cytokines such as interleukins, interferons (alpha., beta, or delta.) and TNF, chlorambucil, busulfan, carmustine, lomustine, semustine, streptozocin, dacarbazine, cytarabine, mercaptopurine, thioguanine, vindesine, etoposide, teniposide, dactinomycin, daunorubicin, doxorubicin, bleomycin, plicamycin, mitomycin, L-asparaginase, hydroxyurea, methylhydrazine, mitotane, tamoxifen, fluoxymesterone, and COX-2 inhibitors.

24. The method of claim 21 wherein the second therapeutic agent is a polypeptide, including soluble forms thereof, selected from the group consisting of Flt3 ligand, CD40 ligand, interleukin-2, interleukin-12, 4-1 BB ligand, anti-4-1 BB antibodies, TRAIL, TNF antagonists and TNF receptor antagonists including TNFR/Fc, Tek antagonists, TWEAK antagonists and TWEAK-R antagonists including TWEAK-R/Fc, VEGF antagonists including anti-VEGF antibodies, VEGF receptor antagonists, CD 148 binding proteins, and nectin-3 antagonists.

25. The method of claim 11 wherein the ADAM disintegrin domain is administered parenterally.

27. The method of claim 26 wherein:

(a) the integrin is α

_vβ

₃ and the ADAM is ADAM-17, ADAM-20, or ADAM-22;

(b) the integrin is α

₂β

₁ and the ADAM is ADAM-23;

(c) the integrin is α

₅β

₁ and the ADAM is ADAM-10, ADAM-21, ADAM-22, or ADAM-23;

(d) the integrin is α

₆β

₁ or α

₆β

₄ and the ADAM is ADAM-10, ADAM-17, ADAM-22, or ADAM-23;

or

(e) the integrin is α

_vβ

₅ and the ADAM is ADAM-10, ADAM-15, or ADAM-23.

29. The method of claim 28 wherein the integrin is present on a cell surface.

30. The method of claim 28 wherein the integrin is selected from the group consisting of α

_vβ

₃, α

₂β

₁, α

₅β

₁, α

₆β

₁, α

₆β

₄, and α

_vβ

₅.

32. The method of claim 30 wherein the ADAM disintegrin domain polypeptide comprises an ADAM disintegrin domain from ADAM-8, ADAM-9. ADAM-10, ADAM-I5, ADAM-17, ADAM-20, ADAM-21, ADAM-2. ADAM-23, or ADAM-29.

33. The method of claim 30 wherein the ADAM disintegrin domain polypeptide comprises an ADAM disintegrin domain from ADAM-1 7, ADAM-20, or ADAM-23.

26. A method for inhibiting the biological activity of an integrin selected from the group consisting of α

_vβ

₃, α

₂β

₁, α

₅β

₁, α

₆β

₁, α

₆β

₄, and α

_vβ

₅ comprising contacting the integrin with an inhibition-effective amount of an ADAM disintegrin domain polypeptide.

28. A method for identifying a compound that modulates integrin biological activity and/or modulates the interaction between an integrin and an ADAM disintegrin domain comprising:

(a) combining a test compound with an integrin and an ADAM disintegrin domain polypeptide that binds to the integrin; and

(b) determining whether the test compound alters the binding of the ADAM disintegrin domain polypeptide to the integrin.

31. A method for identifying a compound that inhibits endothelial cell migration and/or angiogenesis comprising:

(a) combining a test compound with endothelial cells and with an ADAM disintegrin domain polypeptide that binds to endothelial cells; and

(b) determining whether the test compound alters the binding of the ADAM disintegrin domain polypeptide to the endothelial cells.

34. The method of claim 31 wherein the ADAM disintegrin domain polypeptide comprises an ADAM disintegrin domain from ADAM-8, ADAM-9, ADAM-10, ADAM-15, ADAM-17, ADAM-20, ADAM-21, ADAM-22, ADAM-23, or ADAM-29.

35. The method of claim 31 wherein the ADAM disintegrin domain polypeptide comprises an ADAM disintegrin domain from ADAM- 17, ADAM-20, or ADAM-23.